Purification and Characterization of Esterase 6A, a Trimeric Esterase of the House Mouse (Mus musculus)

Abstract

Esterase 6A was isolated from mouse lung and purified 440-fold by ion-exchange chromatography, inverse ammonium sulphate gradient solubilization, gel filtration and isoelectric focusing. The resultant product was apparently homogenous by the criteria of polyacrylamide gel electrophoresis and immunodiffusion, and consisted of the electrophoretic form 6A3. A single species of subunit was present on sodium dodecyl sulphate gel electrophoresis. The molecular weight of the native protein was found to be about 178000 with a subunit molecular weight of about 60000. The equivalent weight obtained by active-site titration with diethyl-p-nitrophenyl phosphate was approximately 178 000 g/mol, indicating a functional asymmetry in the trimer. The enzyme was shown to have a high affinity for 4-nitrophenyl hexanoate (Michaelis constant K , = 4.4 pmol/l) with a relatively low catalytic efficiency (catalytic constant k,,, = 12 s-'). Esterase 6A was immunologically related to esterase 1 and esterase 9, with which it is genetically closely linked. Further properties of the three esterases were compared.

Purification and characterization of a carboxylesterase involved in insecticide resistance from the mosquito Culex quinquefasciatus

“…The carboxylesterase is glycosylated, as shown by the thymol/H2SO4 staining for glycoprotein detection (Gerard, 1990). Glycosylation is characteristic of the vertebrate carboxylesterases (Junge et al, 1974;Hashinotsume et al, 1978;Oehm et al, 1982;Robbi & Beaufay, 1983).…”

Environmental Factors Modulating the Stability and Enzymatic Activity of the Petrotoga mobilis Esterase (PmEst)

“…The molecular mass observed by size exclusion chromatography (SEC) from the purified fraction was ~90 kDa ( Fig 1c ), which suggested that the enzyme PmEst could function as a homo-trimer in aqueous solution. A trimeric state was already observed in the esterases from Pseudomonas putida (with a 33 kDa monomer) [ 32 ] and esterase 6A from Mus musculus (with a 60 kDa subunit) [ 33 ], but also a functional tetramer has been reported for estA [ 34 ].…”

“…A common behavior of esterases is a substrate preference towards short-to medium-length acyl chains (C2–C8) rather than long acyl chains. For example the esterase SAestA, purified from Salinispora arenicola [ 36 ], and esterase 6A show preferences for C6 esters [ 33 ]. In our particular case, there is a strong preference for very short-length chains (C2-C4).…”

Εστερασες Του Ψαριου Trachurus Trachurus. Κατανομη Καθαρισμος Και Φυσικοχημικος Χαρακτηρισμος Των Πολλαπλων Μοριακων Μορφων

“…Εστεράσες έχουν βρεθεί να συμμετέχουν στο μεταβολισμό (α) των λιπιδίων, όπως οι εστεράσες 1 και 2, όπου με το ηλεκτρονικό μικροσκόπιο και μετά από ιστοχημική χρώση διαπιστώθηκε εστερασική ενεργότητα σε λ ι π ιδικά σταγονίδια και πρωτογενή χυλομικρά (Booking and von Deimling, 1982), η εστεράση 6Α (CaE) από πνεύμονες ποντικού (Oehm et al, 1982) και οι Α-εστεράσες που η δράση τους είναι όμοια με εκείνη των φωσφολιπασών C και D (Mackness, 1989), (β) των στεροιδών εστέρων (εστέρες υδροκορτιζόνης) (Hattori et al, 1981, Ali et al, 1985 και (γ) των ακυλιωμένων πρωτεϊνών (Basu et al, 1988). Οι εστεράσες φαίνεται ακόμα ότι συμμετέχουν σε πορείες μεταφοράς ουσιών μέσα στο κύτταρο.…”

“…(α) μονομερές (Bauminger and Levine, 1971, Heymann et al, 1971, Ikeda et al, 1977, Goppinger et al, 1978, Mentlein et al, 1980, Lexow et al, 1980, Otto et al, 1981, Tsujita and Okuda, 1983a, Matsuo et al, 1985, Arav and Rimon, 1987, Korza and Ozols, 1988, Ozols, 1989, Torres et al, 1988, Tsujita et al, 1988, (β) διμερές (Barker and Jencks, 1969, Benohr and Krisch, 1967b, Horgan et al, 1969b, Haugen and Suttie, 1974, Ikeda et al, 1977, Young et al, 1986, Okada and Wakabayashi, 1988, Higa et al, 1989, (γ) τριμερές (Arndt et al, 1973, 1978, Aune, 1973, Mentlein et al, 1980, Oehm et al, 1982, Tsujita et al, 1982, Robby and Beaufay, 1983, Tsujita and Okuda, 1983b, Harano et al, 1988, Lith et al, 1989) και (δ) τετραμερές (Muensh et al, 1976, Lockridge et al, 1979, Rotundo, 1984.…”